Human noroviruses are a leading global cause of severe childhood diarrhea and gastroenteritis outbreaks. However, development of therapeutic and preventative strategies has been severely limited by the inability to propagate these viruses in culture. We recently discovered that noroviruses infect B cells in vitro and in vivo, and that they utilize commensal bacteria as a co-factor for infection. These novel findings led directly to our development of the first human norovirus cultivation system, a tool that will revolutionize norovirus research. We are now uniquely poised to identify host restriction and susceptibility factors influencing human norovirus infection of B cells as well as viral determinants of B cell infection. We hypothesize that this information will reveal key virus-host interactions that can guide the development of novel therapeutics, vaccine candidates, and enhanced diagnostic strategies. In Specific Aim 1, we will utilize complementary targeted and unbiased approaches to develop a comprehensive set of host factors that either restrict or promote human norovirus infection of B cells. In Specific Aim 2, a multi-pronged approach will be undertaken to identify viral determinants of in vitro replicative fitness. Strategies include serial passaging of virus in permissive cells and testing a diverse set of human norovirus genotypes for B cell infectivity. We will also construct a human norovirus reverse genetics system to test adaptive mutations for phenotypic effects. These studies will not only provide critical insight into host and viral determinants of B cell infection by human noroviruses but they should also culminate in the development of 2nd generation culture systems supporting robust infection of a broad panel of viral genotypes.